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Abstract: The frequency of extramedullary infiltration (EMI) in acute myeloblastic leukemia (AML) is reported up to 40% and most prevalent in myelo-monoblastic and monoblastic subtypes of AML (M4 and M5 according to FAB classification). The majority of patients with EMI suffered poor prognosis. To explore mechanism underlying EMI, we analyzed SHI-1 cells, a highly invasive human acute monocytic leukemia cell line, and found their strong expression of matrix metalloproteinase 2 (MMP-2), membrane type 1 MMP (MT1-MMP) and tissue inhibitor of metalloproteinase 2 (TIMP-2). SHI-1 cells showed higher invasive ability to traverse reconstituted basement membranes (Matrigel) and stronger activation of proMMP-2 than other leukemia cell line such as NB4, K562, U937 and THP-1 cells. When co-cultured with bone marrow stromal cells (BMSCs), the invasive capacity and proMMP-2 activation of SHI-1 cells enhanced remarkably. Furthermore, the inhibition of MMP-2, MT1-MMP, or TIMP-2 by small interfering RNA (siRNA) substantially impaired SHI-1 cells invasion and decreased proMMP-2 activation. In the contrast, up-regulated expression of TIMP-2 for 2–3 folds level increased cell invasion and proMMP-2 activation. These results demonstrated that constitutively high expression of MMP-2, MT1-MMP and TIMP-2 in SHI-1 cells facilitated cell invasion by promoting proMMP-2 activation. Moreover, up-regulation of TIMP-2 exhibited not a repressive but an activating effect on SHI-1 cells invasion. Our study indicated that increasing TIMP-2 in AML patients with EMI may potentially cause adverse effects, particularly in patients containing high levels of MMP-2 and MT1-MMP. [Copyright &y& Elsevier]

Abstract: Besides cytopenia, dysplasia is crucial characteristic of MDS-RA. To summarize the morphological features that contribute to the diagnosis of MDS-RA, 48 RA patients with abnormal karyotype were analyzed for the features of morphological and cytogenetical abnormalities and the relationships between them. 46 MDS-RA patients with normal karyotype and 207 patients with non-MDS anemia were enrolled into control groups. More conspicuous and diverse dysplasia can be found in abnormal karyotype MDS-RA than those in control groups (P <0.05). Apparent dysplasia in granulocyte and megakaryocytoid lineages may provide valuable evidence for the diagnosis. Dysplasia occurred more frequently in patients with severe chromosome abnormalities. [Copyright &y& Elsevier]

Abstract: We have studied forty Chinese adult ALL patients at newly diagnosis, using standard primers and protocols of BIOMED-2 multiplex PCR, to determine the feasibility of Ig and TCR gene rearrangements as diagnostic and patient-specific MRD-RQ-PCR targets for molecular monitoring. Clonal IGH, IGK, IGL, TCRB, TCRG and TCRD rearrangements were found in 86%, 22%, 9%, 19%, 77%, 55% of adult patients with B-lineage ALL, respectively. While in T-ALL, clonal IGH, TCRB, TCRG and TCRD rearrangements were detected in 6%, 83%, 78%, 33% of patients. Several specialties in the pattern of Ig/TCR gene rearrangements in Chinese adult ALL patients in comparison with those reported for children and adult patients in other countries have been noted. These results are useful for further MRD-RQ-PCR detection and quantification for all patients. [Copyright &y& Elsevier]

Abstract: CCAAT/enhancer binding proteins (C/EBPs) are a family of transcription factors that have been implicated in diverse cellular functions such as cellular differentiation and proliferation, and inflammatory processes. C/EBPζ, also known as GADD153, CHOP10, and DDIT3 has been found associated with the development of myxoid liposarcoma and the progression of melanoma. To investigate the correlation of C/EBPζ transcript levels with the development of leukemia, samples from 187 patients with myelodysplastic syndrome (MDS), acute myeloid leukemia (AML), and chronic myeloid leukemia (CML) were examined for C/EBPζ mRNA using real-time quantitative PCR (RQ-PCR). RQ-PCR analysis demonstrated the median levels of C/EBPζ were significantly decreased in MDS, AML, and CML patients compared with normal controls (1.40, 0.96, 2.60 versus 14.69, P <0.0001). Significant differences were also observed between patients with CML and with AML or MDS. These results suggest that the insufficient dosage of C/EBPζ might be involved in the development of leukemia. [Copyright &y& Elsevier]

We obtained a full-length cDNA fragment encoding human O6-methylguanine-DNA-methyltransferase (MGMT) from the liver tissue of a patient with cholelithiasis by RT-PCR and confirmed by DNA sequencing. The polycistronic retrovirus vector G1Na-MGMT-Neor-IRES-MDR1 was constructed and verified by restriction endonuclease analysis and DNA sequencing. The vector was transfected into packaging cells GP+E86 and PA317 by the LipofectAMINE method. Cord blood CD34+ cells were transfected with the supernatant of retrovirus containing human MGMT and MDR1cDNA under stimulation of hematopoietic growth factors. PCR, RT-PCR, Southern Blot, Western Blot, FACS and MTT analyses showed that dual drug resistance genes have been integrated into the genomic DNA of cord blood CD34+ cells and expressed efficiently. The transgene cord blood CD34+ cells conferred 5.8–6.3-fold stronger resistance to P-glycoprotein effluxed drugs and 5-fold to BCNU than untransduced cells. The polycistronic retrovirus vector mediated transfer of two different types of drug resistance genes into human cord blood CD34+ cells and co-expression provided an experimental foundation for improving combination chemotherapy tolerance in clinical practice. [Copyright &y& Elsevier]

Abstract: Real-time quantitative reverse transcriptase polymerase chain reaction method was established for detecting the expression levels of WT1 gene and WT1+17AA isoforms in 226 acute myeloid leukemia (AML) bone marrow (BM) cells. The results showed that WT1 gene was 2–3 logarithms expressed more in AML BM cells at initial diagnosis or relapse than in normal BM cells (p <0.001), with predominant WT1+17AA isoforms expression (the ratio of WT1+17AA/WT1 more than 0.50). Interestingly the ratio of WT1+17AA/WT1 was statistically higher in relapsed AMLs than in initially diagnosed (p =0.01), speculating that WT1+17AA isoforms might participate in AML relapse. [Copyright &y& Elsevier]

Background BCL11A encodes a C2H2 type zinc-finger protein. During normal haematopoietic cell differentiation BCL11A expression is down-regulated. Data in mice suggest up-regulation of BCL11A is involved in the pathogenesis of myeloid leukaemias. BCL11A expression in persons with acute myeloid leukaemia (AML) is not systematically studied. Objective Interrogate associations between BCL11A expression at diagnosis and clinical and laboratory valuables and outcomes in newly-diagnosed persons with AML. Methods We determined BCL11A mRNA levels in bone marrow and blood mononuclear cells in 292 consecutive newly-diagnosed subjects with AML by reverse transcript and real-time polymerase chain reaction. Data were compared to mRNA levels in bone marrow cells of normals. Results Subjects with BCL11A transcript levels at diagnosis exceeding the median value of 2.434 (±3.423 SD; 25th–75th inter-quartile range, 1.33–4.29) had higher WBC levels, a greater proportion of bone marrow myeloblasts, were more likely to be FAB M 0 subtype, less likely to be FAB M 3 subtype, more likely to be in the intermediate cytogenetic risk cohort, less likely to have a complex karyotype and more likely to have DNMT3A R882 and FLT3 -ITD mutations than subjects with transcript levels below the median value. In 89 subjects receiving conventional induction chemotherapy the complete remission rate was 54% (95% confidence interval [CI]; 33, 75%) in the lower BCL11A cohort and 65% (45, 85%; P = 0.26) in the higher BCL11A cohort. 3 year survival was 33% (2, 65%) in the lower BCL11A cohort and 15% (0, 39%; P = 0.35) in the high BCL11A cohort. Conclusion BCL11A transcript levels at diagnosis was significantly associated with several clinical and laboratory variables. There were also non-significant associations with complete remission rate and survival. These data suggest a possible role for BCL11A expression in AML biology. [ABSTRACT FROM AUTHOR]